The phenomenon of bacterial resistance through micro-evolutionary change has been well documented. Substantial concerns over the degrading effectiveness of classic broad spectrum antibiotics, such as penicillin and its analogues, as the microbes have adapted new protection mechanisms, has led to medical protocols that attempt to minimize their usage. The goal is to delay the time when larger segments of the population may find the effectiveness of critically needed antibiotics compromised for their needs. To delay this problem, such powerful anti-microbial agents as Vancomycin are prescribed only under the most stringent use protocols. In addition, there are few pharmacological compounds that have been found to be effective for treatment of viruses, and the few that exist are very strain-specific.
The pharmaceutical industry is desperately seeking new ways to battle infection. The myriad attempts to develop improved types of antibotics and anti-viral agents include mechanisms ranging from various innovative chemical inactivation mechanisms, gene-based techniques of drug production, and tailored anti-infective medications for individual patients. What all of these approaches have in common is that they are chemical: drugs that are ingested and then act biochemically on the host. The disadvantage is that, while the pathogens may be destroyed, the patient may also be compromised or worse, due to drug toxicity.
A further problem is that, in general for a severe systemic infection, there still exist antibiotic drugs that can effectively destroy bacterial pathogens. However, what often kills the patient are the toxic poisons that are released by the bacteria as they are lysed (destroyed) due to either immune system action or antibiotic effects. There are two categories of such substances: endotoxins, lipopolysaccharides that are associated with the cellular walls of gram-negative bacteria, and exotoxins, soluble proteins which are diffusible and may act at different sites from the bacterial invasion as some of the most potent poisons known, comparable to strychnine and snake venom. It may help the patient very little to destroy a bacterium that leaves a parting gift of poison for the host as it dies. A treatment is needed that has the potential to both destroy bacteria in biological fluids such as blood, and also allow toxins to be filtered out of the blood before they can injure the patient. This phenomenon is applicable to both humans and animals.
Various methods of non-chemical purification of biological fluids, medicines, vaccines, etc., have been proposed to destroy or inactivate pathogens, including bacteria, viruses, and fungi, in the liquids. These methods typically generate heat during the purification process and may introduce impurities depending on the process. This heat can easily damage the active ingredients or critical live tissues that perform the essential functions of the fluids. In the case of biological fluids, if these tissues are thermally processed, they may become non-functional, as in the case of some clotting compounds, or vital living cells such as erythrocytes (red blood cells, RBC) or leukocytes (white blood cells, WBC) may be altered or damaged in ways that both mar instant function or compromise cellular stability, hardiness, or reproduction. When treating delicate biological fluids such as blood, these processes have had marginal success because blood functions properly in a very narrow temperature range around the normal 37 degrees C. (nominally 98.6 degrees F.).
Blood that is drawn from donors and banked for transfusion currently must go through multiple exhaustive steps to ensure safety for use, and there are large volumes of scarce blood and plasma that must be destroyed when they are found to be contaminated. A pasteurization technology that is able to cycle fluids such as blood through an apparatus outside the body to destroy the bacterial infection, and then filter out the toxins released in the process, and permit blood or plasma to be safely used rather than wasted will provide significant benefit to the health care system. Such a safe and efficacious technology should also permit cost-saving protocol changes in the collection and handling of biological fluids such as blood.
A number of minimal thermal processes have been developed for some of these applications, including ultra-filtration, ozonation, pulsed ultraviolet light, irradiation, high hydrostatic pressure (HHP) and pulsed electric field (PEF) discharge.
PEF discharge has been shown to be very effective for killing bacteria within medically useful liquids that are not subject to degradation, such as vaccines, medications, and other sterile substances. PEF discharge is considered to be one of the premier new technologies with a great potential of replacing thermal, chemical and other pasteurization and sterilization technologies. However, there are a number of drawbacks of the PEF discharge technology. For example, ohmic heating still occurs during the PEF discharge, which causes the temperature of the liquid being treated to rise. Hence, a cooling system must be used in order to maintain the liquid at a low temperature. A significant amount of energy is wasted with unwanted heating and cooling of the liquid. Also, the requirement of a cooling system adversely increases the time required to treat the liquid. In addition, the PEF electrodes are immersed directly in the liquid. Since the electrodes contact the liquid, they are regarded as a major contamination source to the liquid due to oxidation of the electrodes during discharge. The electrodes must therefore be replaced regularly, which increases maintenance time and costs.
With respect to ozonation, numerous research reports have demonstrated the antiviral effect of ozone exposure. For example, K. H. Wells, J. Latino, J. Gavalchin and B. J. Poiesz, Blood, 78, 1882–90 (1991), reported more than 11 logs inactivation of HIV-1 virus in human blood that was exposed to ozone at a concentration of 1,200 ppm for two hours. J. M. Vaughn, Y. S. Chen, K. Lindburg and D. Morales, Appl. Environ. Microbiol., 53, 2218–21 (1987), reported the use of ozone to inactivate simian rotavirus SA-11 and human rotavirus type 2 (Wa) at 4° C. by using single-particle virus stocks, and found that although the human strain was clearly more sensitive; both virus types were rapidly inactivated by ozone concentrations of 0.25 mg/liter or greater at all pH levels tested.
Use of ozone to kill viruses in blood and blood products has received increasing attention in the medical field, due to its high effectiveness, cost efficiency, and simplicity, with minimal collateral damage to blood cellular metabolism. Ozone is a strong oxidative, and can react with blood to form compounds which are identical to those produced by a human's own immune system to destroy viruses and bacteria. Some of these compounds include oxygen atoms, hydrogen peroxide, and lipid peroxide. Research in the Wells et al. article has indicated that the antiviral effects of ozone include viral particle disruption, reverse transcriptase inactivation, and/or a perturbation of the ability of the virus to bind to its receptor on target cells. Based on a study on the mechanism of enteroviral inactivation by ozone with poliovirus 1 as the model virus, D. Roy, P. K. Wong, R. S. Engelbrecht and E. S. Chian, Appl. Environ. Microbiol., 41, 718–23 (1981) concluded that the damage to the viral nucleic acid is the major cause of poliovirus 1 inactivation by ozone.
In addition, use of ozone is entering commercial practice for purification of bottled water, but requires strict limitation of its concentration because of concerns over potential formation of the suspected carcinogen bromate when excess ozone concentration is permitted to interact with minerals in the water. This becomes an even greater concern when treating biological fluids, such as blood products since ozone can dissolve in the water of the blood plasma and therefore remain for an extended period of time. With biological fluids, the ozone must therefore be “inactivated” through the passage of time or by some other method. The lingering ozone may be an advantage for purifying bottled water, but in biological fluids, control of all biochemical process is critical.
Thus, while ozone and other minimally thermal methods are being researched for treatment of biological fluids, improved methods are desired for treating biological fluids such as blood without degrading their natural characteristics or generating toxic byproducts.